Persistent pulmonary hypertension of the newborn (PPHN) is a clinical syndrome characterized by the inability to achieve or sustain the normal decline in pulmonary vascular resistance (PVR) at birth. Insight into basic mechanisms which maintain high PVR in the normal fetus and contribute to the normal or unsuccessful transition of the pulmonary circulation at birth will lead to greater understanding of the pathogenesis, pathophysiology and treatment of PPHN. Although associated with diverse neonatal cardiopulmonary disorders, common pathophysiologic features of PPHN include pulmonary hypertension, abnormal vasoreactivity, and vascular remodeling. Recent studies from several laboratories have demonstrated the important role of the endothelium in control of vascular tone, reactivity and growth, and that impaired function may contribute to the pathophysiology of several clinical disorders. Little is known, however, about the role of the endothelial cell in regulating vascular function in the developing lung. Endothelium-derived relaxing factor or nitric oxide (EDRF/NO) has recently been shown to contribute to vasoregulation in the ovine fetal and transitional pulmonary circulations. In addition, preliminary studies suggest that inhaled NO may be effective in the clinical treatment of PPHN, reducing the need for extracorporeal membrane oxygenation therapy. Little is known, however, about maturation- related changes in EDRF/NO activity, its physiologic roles in the perinatal lung, mechanisms regulating its activity, whether its activity is altered in severe neonatal pulmonary hypertension, and potential mechanisms contributing to abnormal EDRF/NO function. Based on past studies and preliminary data, we hypothesize that 1) EDRF/NO activity progressively increases during late-gestation in the fetal lung; 2) EDRF/NO activity is augmented with postnatal maturation; and 3) chronic intrauterine pulmonary hypertension decreases EDRF/NO activity, causing abnormal pulmonary vasoreactivity and failure of postnatal adaptation. Therefore, the goals of this proposal are 1) to characterize mechanisms underlying normal developmental changes in EDRF/NO activity; 2) to determine the role of EDRF/NO activity in normal perinatal physiology; and 3) to characterize mechanisms underlying altered EDRF/NO activity with vascular injury in a model of chronic intrauterine pulmonary hypertension. To address these questions, we propose a series of parallel in vivo and in vitro experiments, examining hemodynamic responses to pharmacologic and physiologic stimuli in chronically-prepared fetal lambs and small pulmonary arteries in smooth muscle bath preparations, along with biochemical, molecular and immunostaining analyses of Type III nitric oxide synthase and soluble guanylate cyclase in the normal and hypertensive fetal, neonatal and adult pulmonary circulations in sheep.